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Mechanism Underlying Hippocampal Long-Term Potentiation And www.nature.com/scientificreports OPEN Mechanism underlying hippocampal long‑term potentiation and depression based on competition between endocytosis and exocytosis of AMPA receptors Tomonari Sumi1,2* & Kouji Harada3 n‑methyl‑D‑aspartate (NMDA) receptor‑dependent long‑term potentiation (LTP) and long‑term depression (LTD) of signal transmission form neural circuits and thus are thought to underlie learning and memory. These mechanisms are mediated by AMPA receptor (AMPAR) trafcking in postsynaptic neurons. However, the regulatory mechanism of bidirectional plasticity at excitatory synapses remains unclear. We present a network model of AMPAR trafcking for adult hippocampal pyramidal neurons, which reproduces both LTP and LTD. We show that the induction of both LTP and LTD is regulated by the competition between exocytosis and endocytosis of AMPARs, which are mediated by the calcium-sensors synaptotagmin 1/7 (Syt1/7) and protein interacting with C-kinase 1 (PICK1), respectively. Our result indicates that recycling endosomes containing AMPAR are always ready for Syt1/7-dependent exocytosis of AMPAR at peri-synaptic/synaptic membranes. This is because molecular motor myosin Vb constitutively transports the recycling endosome toward the membrane in a Ca2+‑independent manner. Synaptic plasticity is generally regulated by the release of various neurotransmitters from the presynaptic mem- brane and/or by varying the density, types, and properties of neurotransmitter receptors at the postsynaptic membrane. NMDA (N-methyl-D-aspartate) receptor-dependent long-term potentiation (LTP) and long-term depression (LTD) of signal transmission in excitatory neurons, such as hippocampal pyramidal neurons, is thought to underlie the formation of neuronal circuits during learning and memory 1–3. Fast excitatory neuro- transmission in the mammalian brain is predominantly mediated by the AMPA (α-amino-3-hydroxy-5-methyl- 4-isoxazolepropionic acid) receptor (AMPAR) at the postsynaptic membrane. AMPAR is tetrameric ion channel composed of the subunits GluA1–A4 (or named as GluR1–R4). In hippocampal pyramidal neurons, the GluA1/ A2 heterotetramer (Fig. 1a) is the most dominant AMPAR subtype, followed by the GluA2/A3 heterotetramer 4. It is well established that AMPAR trafcking in postsynaptic neurons plays a decisive role in the induction of LTP and LTD 5–8, whereas the dominant pathway and regulatory mechanism for AMPAR trafcking remains a controversial issue. NMDA receptor-dependent LTP and LTD are triggered by standard high-frequency stimulations (e.g., one or more trains of 100 Hz stimulation)9,10 and low-frequency stimulations (LFS; e.g., 700–900 pulses at 1 Hz)11–14, respectively. It is widely accepted that a high rise and lower rise in intracellular Ca 2+ concentrations mediated by NMDA receptor activation are required for the expression of LTP and LTD, respectively. Nevertheless, it 1Research Institute for Interdisciplinary Science, Okayama University, 3-1-1 Tsushima-Naka, Kita-ku, Okayama 700-8530, Japan. 2Department of Chemistry, Faculty of Science, Okayama University, 3-1-1 Tsushima-Naka, Kita-ku, Okayama 700-8530, Japan. 3Department of Computer Science and Engineering, Toyohashi University of Technology, 1-1 Hibarigaoka, Tempaku-cho, Toyohashi, Aichi 441-8580, Japan. *email: [email protected] SCIENTIFIC REPORTS | (2020) 10:14711 | https://doi.org/10.1038/s41598-020-71528-3 1 Vol.:(0123456789) www.nature.com/scientificreports/ Figure 1. AMPAR trafcking model at hippocampal postsynaptic neurons. (a) AMPAR which consists of two GluA1 (GluR1) and two GluA2 (GluR2) subunits is the most predominant AMPAR subtype in hippocampal neurons4. Te serine-845 site (S845) of GluA1 is phosphorylated and dephosphorylated by protein kinase A (PKA)35 and protein phosphatase 2B (PP2B, or Calcineurin, CaN)36, respectively. Te serine-880 site (S880) of GluA2 is phosphorylated and dephosphorylated by protein kinase C (PKC)37 and protein phosphatase 2 (PP2A), respectively. (b) A-kinase anchoring protein 150 (AKAP150)34,43,44 is an anchoring protein that organizes PKA, PP2B, and PKC for phosphoregulation of AMPARs at the synaptic membrane, and thus acts as the AKAP signaling complex. Te AKAP signaling complex forms the dimer as shown in (b) (though, for simplifcation, not shown in (d–e)). (c–h) Experimentally characterized elementary processes involved in the AMPAR trafcking cycle at a hippocampal postsynaptic neuron. (c) A model of tethering the GluA1 and GluA2 subunits to the AKAP150 signaling complex at the synaptic membrane through SAP97 and GRIP1, respectively34,43,44. (d) A model of phosphorylation and dephosphorylation reactions of the AMPAR due to Ca 2+ signaling. Dephosphorylation of the S845 site of GluA1 is caused by PP2B 36, and SAP97 dissociates from GluA1. Te S880 site of GluA2 is phosphorylated by PKC, and PICK1 binds to the GluA2 subunit instead of GRIP1 37,38. (e,f) An endocytic model for synaptic vesicles containing AMPAR mediated by the calcium-sensor PICK1 37,38,47. 40–42 (g) Active transport of the recycling endosomes by molecular motor myosin V b . In the recycling endosome, PP2A causes the dephosphorylation of S880 of GluA2, and GRIP1 instead of PICK1 binds to GluA2 46. (h) An exocytic model of the recycling endosome triggered by Ca 2+-sensor synaptic vesicle protein synaptotagmin 1 (Syt1) together with synaptotagmin 7 (not shown) and synaptobrevin-2 (Syb2)/VAMP2, complexin (not shown), amongst others22,23,48–50,61. has been reported that a channel blocker for Ca 2+ infux through the NMDA receptor does not block LTD in the hippocampal neurons of immature rodents, thereby suggesting that LTD is attributed to an ion-channel- independent, metabotropic form of NMDA-receptor signaling15. Subsequently, it was carefully reexamined how varying extracellular Ca2+ levels and blocking NMDA-receptor channel ion fux with the same channel blocker afected the induction of LTD. It was reconfrmed that the LTD induced by a standard LFS in hippocampal neu- rons of both adult and immature rodents was dependent on ionotropic NMDA-receptor signaling 16. For more than 20 years, it has been known that Ca 2+/calmodulin-dependent protein kinase (CaMKII) activity is necessary for LTP. Ca2+ ions incorporate into the cell through NMDA receptor and bind to calmodulin (CaM). Consequently Ca2+/CaM is bound to CaMKII. If CaMKII is knocked out, Ca 2+/CaM binds directly to NMDA receptors, resulting in inactivation of the NMDA receptor channel and an accordingly signifcantly lower rise in Ca2+ concentrations17,18. On the other hand, if Ca2+/CaM-bound CaMKII binds to the NMDA receptor and inhibits the direct binding of Ca2+/CaM to the NMDA receptor, this maintains the channel activity of the NMDA SCIENTIFIC REPORTS | (2020) 10:14711 | https://doi.org/10.1038/s41598-020-71528-3 2 Vol:.(1234567890) www.nature.com/scientificreports/ receptor. Tis is one of the most important CaMKII functions at the early stage of NMDA receptor-dependent LTP induction process. In addition, the Ca 2+/CaM-CaMKII bound to the NMDA receptor phosphorylates GluA119, resulting in increased channel conductance of AMPAR20. Te activation of AMPAR by CaMKII also plays a crucial role in the induction of LTP. In the present study, we build a network model to reproduce NMDA- receptor dependent bidirectional synaptic plasticity of hippocampal neurons. All of the molecular mechanisms incorporated into the model work downstream of these CaMKII functions; thus, the normal activities of CaMKII are necessary and implicitly assumed in our network model. In fact, NMDA receptor-dependent Ca 2+-infux is introduced in the network model as the input for LTP and LTD simulations. In addition, it is assumed that the strength of LTP and LTD induction is in proportion to the change in the AMPAR population on the postsynaptic membrane. Tus, the efects of GluA2-lacking Ca 2+-permeable AMPARs are not taken into consideration in our simulations. However, this assumption does not infuence the prediction of LTP expression by our network model, because it has been demonstrated that LTP in the hippocampal CA1 region does not require insertion or activation of GluA2-lacking AMPARs 21. NMDA-receptor dependent LTP is thought to occur by incorporating AMPARs into the synaptic membrane through either the exocytosis of the recycling endosome at peri-synaptic/synaptic membranes22–25, the cell surface long-range lateral difusion of AMPAR from the dendrites/extrasynaptic region26–29, or a combination of the two. Recently, Penn et al. examined the efects of crosslinking immobilization of pre-existing membrane AMPARs on early LTP and observed slow continuous development of early LTP without short-term potentiation (STP)26. Tey also observed that blocking exocytosis of AMPARs completely abolished early LTP and caused only STP induction26. Te former and latter respectively indicate the following: (1) lateral difusion movement of AMPARs exocytosed at the synaptic/peri-synaptic membranes as well as the extrasynaptic membranes is signifcantly suppressed due to obstacle efects arising from jamming/crowding of crosslinked AMPARs, as predicted by computer simulations30; (2) although the recruitment of pre-existing membrane AMPARs from the peri-synaptic membrane works for the induction of STP, it is insufcient for maintaining the expression of early LTP. Terefore, the difusion dynamics on the membrane strongly afect the form of LTP expression and play a crucial role in the rapid short-range difusion relocation of AMPARs from the peri-synaptic to
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